The treatment of neuropathic pain continues to challenge clinicians, in part because of limited efficacy of available treatments and a lack of pain-specific drug targets. Mas-related G-protein-coupled receptors (Mrgprs) represent a novel family of G-protein-coupled receptors (GPCRs) that are specifically expressed in small-diameter sensory neurons. Our preliminary work suggests that some Mrgprs, in particular MrgprC, may constitute a novel inhibitory system for persistent pain. Intrathecal administration of a peptide (BAM 8-22) and a small molecule non-peptide MrgprC agonist (compound 58) both attenuated neuropathic pain in rats and in wild type mice. However, BAM 8-22 lost this analgesic action on neuropathic pain in Mrpgr mutant mice. Using complementary behavioral, electrophysiological, and molecular biological approaches, we aim to assess the therapeutic value of MrgprC as a novel target in the treatment of neuropathic pain, and importantly, to examine the cellular and molecular mechanisms underlying MrgprC-mediated pain inhibition. Specifically, Aim 1 will characterize in detail the effects of intrathecal administration of the two MrgprC agonists on tactile allodynia, mechanical hyperalgesia, heat hyperalgesia and spontaneous pain in rats after an L5 spinal nerve injury. We will then examine whether MrgprC siRNA treatment reduces the analgesic efficacy of BAM 8-22. We will also investigate whether an early intervention with BAM 8-22 prolongs attenuation of neuropathic pain, enhances morphine analgesia in neuropathic pain, and prevents postsurgical pain in rats. In Aim 2, we will employ in vitro electrophysiological and molecular biological approaches to study the cellular and molecular mechanisms underlying MrgprC-mediated pain inhibition in the dorsal root ganglion (DRG) neurons. We will use both loss-of-function and gain-of-function strategies to determine if MrgprC activation attenuates high-voltage-gated (HVA) calcium current in DRG cells. We will further determine whether nerve injury up regulates the expression and function of MrgprC in uninjured DRG neurons. In Aim 3 we will study the neurophysiologic basis for MrgprC-mediated pain inhibition in the central nervous system. We will conduct in vivo electrophysiological studies to determine if MrgprC agonist attenuates dorsal horn neuronal sensitization to repetitive noxious inputs and normalizes the established neuronal hyperexcitability induced by the nerve injury. We will investigate whether nerve injury alters the expression of BAM 22, an endogenous Mrgpr ligand, in spinal cord and in DRGs. We postulate that MrgprC agonists may function as anti-hyperalgesic agents during the neuropathic pain state. Because MrgprC is an ortholog to the human MrgprX1, our findings may have important implications for developing new drug leads and mechanism-based treatment strategies for managing neuropathic pain with few side effects.